Deletion of the PDZ₂ but not the PDZ₃ domain of PSD-95 prevents MIS formation. (A, top) Contour representation of a dendritic segment reconstructed from a PDZ₃ mutant PSD-95–overexpressing cell illustrating the preservation of numerous MISs (numbers refer to presynaptic terminals [*] making synaptic contacts on the spine). (bottom) Three-dimensionally reconstructed segment expressing PDZ₃ mutant PSD-95 illustrating large, complex spines. (B, top) Same as in A (top) but for a PDZ₂ mutant PSD-95. Note the absence of MISs. (bottom) Same as in A (bottom) but for a cell expressing PDZ₂ mutant PSD-95. MISs are absent, but large spines are still present. (C) Quantitative analysis of the proportion of MISs observed under control conditions (ctrl; n = 8 cells; 145 spines) and in cells transfected with EGFP (n = 4 cells; 164 spines), PSD-95 (n = 7 cells; 234 spines), PDZ₃ mutant PSD-95 (n = 3 cells; 86 spines), and PDZ₂ mutant PSD-95 (n = 3 cells; 56 spines; *, P < 0.05 vs. control and PDZ₂). (D) Same as in C but for the spine volume. Expression of PDZ mutants of PSD-95 does not prevent PSD-95–induced spine enlargement (*, P < 0.05 vs. control). (E) Same as in C but for the total PSD area. Expression of PDZ mutants of PSD-95 does not reduce or only partially reduces the increase in PSD area (*, P < 0.05 vs. control). Data are mean ± SEM (error bars). Bars: (A and B, top) 5 μm; (A and B, bottom) 1 μm.